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Wiley Interdisciplinary Reviews.... Nov 2021The 3D organization of the genome facilitates gene regulation, replication, and repair, making it a key feature of genomic function and one that remains to be properly... (Review)
Review
The 3D organization of the genome facilitates gene regulation, replication, and repair, making it a key feature of genomic function and one that remains to be properly understood. Over the past two decades, a variety of chromosome conformation capture (3C) methods have delineated genome folding from megabase-scale compartments and topologically associating domains (TADs) down to kilobase-scale enhancer-promoter interactions. Understanding the functional role of each layer of genome organization is a gateway to understanding cell state, development, and disease. Here, we discuss the evolution of 3C-based technologies for mapping 3D genome organization. We focus on genomics methods and provide a historical account of the development from 3C to Hi-C. We also discuss ChIP-based techniques that focus on 3D genome organization mediated by specific proteins, capture-based methods that focus on particular regions or regulatory elements, 3C-orthogonal methods that do not rely on restriction digestion and proximity ligation, and methods for mapping the DNA-RNA and RNA-RNA interactomes. We consider the biological discoveries that have come from these methods, examine the mechanistic contributions of CTCF, cohesin, and loop extrusion to genomic folding, and detail the 3D genome field's current understanding of nuclear architecture. Finally, we give special consideration to Micro-C as an emerging frontier in chromosome conformation capture and discuss recent Micro-C findings uncovering fine-scale chromatin organization in unprecedented detail. This article is categorized under: Gene Expression and Transcriptional Hierarchies > Regulatory Mechanisms Gene Expression and Transcriptional Hierarchies > Gene Networks and Genomics.
Topics: Cell Nucleus; Chromatin; Chromosomes; Genome; Promoter Regions, Genetic
PubMed: 32987449
DOI: 10.1002/wdev.395 -
Nature Reviews. Immunology Jan 2021
PubMed: 33262449
DOI: 10.1038/s41577-020-00482-y -
Vavilovskii Zhurnal Genetiki I Selektsii Oct 2021Random transgene integration is a powerful tool for developing new genome-wide screening approaches. These techniques have already been used for functional gene...
Random transgene integration is a powerful tool for developing new genome-wide screening approaches. These techniques have already been used for functional gene annotation by transposon-insertion sequencing, for identif ication of transcription factor binding sites and regulatory sequences, and for dissecting chromatin position effects. Precise localization of transgenes and accurate artifact f iltration are essential for this type of method. To date, many mapping assays have been developed, including Inverse-PCR, TLA, LAM-PCR, and splinkerette PCR. However, none of them is able to ensure localization of both transgene's f lanking regions simultaneously, which would be necessary for some applications. Here we proposed a cheap and simple NGS-based approach that overcomes this limitation. The developed assay requires using intentionally designed vectors that lack recognition sites of one or a set of restriction enzymes used for DNA fragmentation. By looping and sequencing these DNA fragments, we obtain special data that allows us to link the two f lanking regions of the transposon. This can be useful for precise insertion mapping and for screening approaches in the f ield of chromosome engineering, where chromosomal recombination events between transgenes occur in a cell population. To demonstrate the method's feasibility, we applied it for mapping SB transposon integration in the human HAP1 cell line. Our technique allowed us to eff iciently localize genomic transposon integrations, which was conf irmed via PCR analysis. For practical application of this approach, we proposed a set of recommendations and a normalization strategy. The developed method can be used for multiplex transgene localization and detection of rearrangements between them.
PubMed: 34755021
DOI: 10.18699/VJ21.068 -
Molecular Phylogenetics and Evolution Aug 2021The advent of high throughput sequencing technologies provides an opportunity to resolve phylogenetic relationships among closely related species. By incorporating...
The advent of high throughput sequencing technologies provides an opportunity to resolve phylogenetic relationships among closely related species. By incorporating hundreds to thousands of unlinked loci and single nucleotide polymorphisms (SNPs), phylogenomic analyses have a far greater potential to resolve species boundaries than approaches that rely on only a few markers. Scleractinian taxa have proved challenging to identify using traditional morphological approaches and many groups lack an adequate set of molecular markers to investigate their phylogenies. Here, we examine the potential of Restriction-site Associated DNA sequencing (RADseq) to investigate phylogenetic relationships and species limits within the scleractinian coral genus Porites. A total of 126 colonies were collected from 16 localities in the seas surrounding the Arabian Peninsula and ascribed to 12 nominal and two unknown species based on their morphology. Reference mapping was used to retrieve and compare nearly complete mitochondrial genomes, ribosomal DNA, and histone loci. De novo assembly and reference mapping to the P. lobata coral transcriptome were compared and used to obtain thousands of genome-wide loci and SNPs. A suite of species discovery methods (phylogenetic, ordination, and clustering analyses) and species delimitation approaches (coalescent-based, species tree, and Bayesian Factor delimitation) suggested the presence of eight molecular lineages, one of which included six morphospecies. Our phylogenomic approach provided a fully supported phylogeny of Porites from the Arabian Peninsula, suggesting the power of RADseq data to solve the species delineation problem in this speciose coral genus.
Topics: Animals; Anthozoa; Arabia; Bayes Theorem; DNA, Ribosomal; Genome, Mitochondrial; Phylogeny; Sequence Analysis, DNA
PubMed: 33813021
DOI: 10.1016/j.ympev.2021.107173 -
Frontiers in Genetics 2022It is now evident that DNA forms an organized nuclear architecture, which is essential to maintain the structural and functional integrity of the genome. Chromatin...
It is now evident that DNA forms an organized nuclear architecture, which is essential to maintain the structural and functional integrity of the genome. Chromatin organization can be systematically studied due to the recent boom in chromosome conformation capture technologies (e.g., 3C and its successors 4C, 5C and Hi-C), which is accompanied by the development of computational pipelines to identify biologically meaningful chromatin contacts in such data. However, not all tools are applicable to all experimental designs and all structural features. Capture Hi-C (CHi-C) is a method that uses an intermediate hybridization step to target and select predefined regions of interest in a Hi-C library, thereby increasing effective sequencing depth for those regions. It allows researchers to investigate fine chromatin structures at high resolution, for instance promoter-enhancer loops, but it introduces additional biases with the capture step, and therefore requires specialized pipelines. Here, we compare multiple analytical pipelines for CHi-C data analysis. We consider the effect of retaining multi-mapping reads and compare the efficiency of different statistical approaches in both identifying reproducible interactions and determining biologically significant interactions. At restriction fragment level resolution, the number of multi-mapping reads that could be rescued was negligible. The number of identified interactions varied widely, depending on the analytical method, indicating large differences in type I and type II error rates. The optimal pipeline depends on the project-specific tolerance level of false positive and false negative chromatin contacts.
PubMed: 35198004
DOI: 10.3389/fgene.2022.786501 -
Molecular Metabolism Mar 2023Oligodendrocyte progenitor cell differentiation is regulated by nutritional signals in the adult median eminence (ME), but the consequences on local myelination are...
OBJECTIVE
Oligodendrocyte progenitor cell differentiation is regulated by nutritional signals in the adult median eminence (ME), but the consequences on local myelination are unknown. The aim of this study was to characterize myelin plasticity in the ME of adult mice in health or in response to chronic nutritional challenge and determine its relevance to the regulation of energy balance.
METHODS
We assessed new oligodendrocyte (OL) and myelin generation and stability in the ME of healthy adult male mice using bromodeoxyuridine labelling and genetic fate mapping tools. We evaluated the contribution of microglia to ME myelin plasticity in PLX5622-treated C57BL/6J mice and in Pdgfra-Cre/ER;R26R-eYFP;Myrf mice, where adult oligodendrogenesis is blunted. Next, we investigated how high-fat feeding or caloric restriction impact ME OL lineage progression and myelination. Finally, we characterized the functional relevance of adult oligodendrogenesis on energy balance regulation.
RESULTS
We show that myelinating OLs are continuously and rapidly generated in the adult ME. Paradoxically, OL number and myelin amounts remain remarkably stable in the adult ME. In fact, the high rate of new OL and myelin generation in the ME is offset by continuous turnover of both. We show that microglia are required for continuous OL and myelin production, and that ME myelin plasticity regulates the recruitment of local immune cells. Finally, we provide evidence that ME myelination is regulated by the body's energetic status and demonstrate that ME OL and myelin plasticity are required for the regulation of energy balance and hypothalamic leptin sensitivity.
CONCLUSIONS
This study identifies a new mechanism modulating leptin sensitivity and the central control of energy balance and uncovers a previously unappreciated form of structural plasticity in the ME.
Topics: Mice; Male; Animals; Myelin Sheath; Leptin; Mice, Transgenic; Median Eminence; Mice, Inbred C57BL
PubMed: 36739968
DOI: 10.1016/j.molmet.2023.101690 -
World Journal of Emergency Surgery :... Oct 2023The creation of an ileostomy or colostomy is a common surgical event, both in elective and in emergency context. The main aim of stoma creation is to prevent... (Review)
Review
BACKGROUND
The creation of an ileostomy or colostomy is a common surgical event, both in elective and in emergency context. The main aim of stoma creation is to prevent postoperative complications, such as the anastomotic leak. However, stoma-related complications can also occur and their morbidity is not negligible, with a rate from 20 to 70%. Most stomal complications are managed conservatively, but, when this approach is not resolutive, surgical treatment becomes necessary. The aim of this mapping review is to get a comprehensive overview on the incidence, the risk factors, and the management of the main early and late ostomy complications: stoma necrosis, mucocutaneous separation, stoma retraction, stoma prolapse, parastomal hernia, stoma stenosis, and stoma bleeding.
MATERIAL AND METHODS
A complete literature research in principal databases (PUBMED, EMBASE, SCOPUS and COCHRANE) was performed by Multidisciplinary Italian Study group for STOmas (MISSTO) for each topic, with no language restriction and limited to the years 2011-2021. An international expert panel, from MISSTO and World Society of Emergency Surgery (WSES), subsequently reviewed the different issues, endorsed the project, and approved the final manuscript.
CONCLUSION
Stoma-related complications are common and require a step-up management, from conservative stoma care to surgical stoma revision. A study of literature evidence in clinical practice for stoma creation and an improved management of stoma-related complications could significantly increase the quality of life of patients with ostomy. Solid evidence from the literature about the correct management is lacking, and an international consensus is needed to draw up new guidelines on this subject.
Topics: Humans; Quality of Life; Ostomy; Surgical Stomas; Colostomy; Ileostomy
PubMed: 37817218
DOI: 10.1186/s13017-023-00516-5 -
Frontiers in Nutrition 2021Healthcare systems worldwide are seriously challenged by a rising prevalence of neurodegenerative diseases (NDDs), which mostly, but not exclusively, affect the... (Review)
Review
Healthcare systems worldwide are seriously challenged by a rising prevalence of neurodegenerative diseases (NDDs), which mostly, but not exclusively, affect the ever-growing population of the elderly. The most known neurodegenerative diseases are Alzheimer's (AD) and Parkinson's disease, multiple sclerosis, and amyotrophic lateral sclerosis, but some viral infections of the brain and traumatic brain injury may also cause NDD. Typical for NDD are the malfunctioning of neurons and their irreversible loss, which often progress irreversibly to dementia and ultimately to death. Numerous factors are involved in the pathogenesis of NDD: genetic variability, epigenetic changes, extent of oxidative/nitrosative stress, mitochondrial dysfunction, and DNA damage. The complex interplay of all the above-mentioned factors may be a fingerprint of neurodegeneration, with different diseases being affected to different extents by particular factors. There is a voluminous body of evidence showing the benefits of regular exercise to brain health and cognitive functions. Moreover, the importance of a healthy diet, balanced in macro- and micro-nutrients, in preventing neurodegeneration and slowing down a progression to full-blown disease is evident. Individuals affected by NDD almost inevitably have low-grade inflammation and anomalies in lipid metabolism. Metabolic and lipid profiles in NDD can be improved by the Mediterranean diet. Many studies have associated the Mediterranean diet with a decreased risk of dementia and AD, but a cause-and-effect relationship has not been deduced. Studies with caloric restriction showed neuroprotective effects in animal models, but the results in humans are inconsistent. The pathologies of NDD are complex and there is a great inter-individual (epi)genetic variance within any population. Furthermore, the gut microbiome, being deeply involved in nutrient uptake and lipid metabolism, also represents a pillar of the gut microbiome-brain axis and is linked with the pathogenesis of NDD. Numerous studies on the role of different micronutrients (omega-3 fatty acids, bioactive polyphenols from fruit and medicinal plants) in the prevention, prediction, and treatment of NDD have been conducted, but we are still far away from a personalized diet plan for individual NDD patients. For this to be realized, large-scale cohorts that would include the precise monitoring of food intake, mapping of genetic variants, epigenetic data, microbiome studies, and metabolome, lipidome, and transcriptome data are needed.
PubMed: 34422879
DOI: 10.3389/fnut.2021.688086 -
Globalization and Health Sep 2021Substandard and falsified (SF) medical products are a global public health threat. The presence and spread of SF drugs negatively affect (1) patients' safety and health...
OBJECTIVE
Substandard and falsified (SF) medical products are a global public health threat. The presence and spread of SF drugs negatively affect (1) patients' safety and health outcomes, (2) national economy, (3) public trust in the healthcare system, and (4) the international fight against serious health challenges such as malaria and antimicrobial resistance. The objective of the current study was to investigate and provide a snapshot analysis of the evolution and developmental patterns of global research publications on SF products.
METHODS
A bibliometric approach was adopted using terms such as fake, falsified, counterfeit, substandard, and others. No language restriction was made. The study period was from 1900 up to 2020. The search strategy was validated and implemented using Scopus database.
RESULTS
The search strategy retrieved 978 documents authored by 2861 researchers from 100 different countries and published in 421 different journals. The retrieved documents received 11,237 citations (11.5 citations per document) with an H-index of 53. The 978 documents retrieved from Scopus were published from 1961 to 2020, giving an average of 16.6 publications per year. The present study indicated that research on SF medical products: (a) has experienced a steep growth from 2001 to 2012 followed by a steady-state growth; (b) was disseminated in a wide range of journals, mainly in the fields of the pharmaceutical industry, analytical chemistry, public health, infectious diseases, and internal medicine; (c) was published by scholars with diverse and distant geographical backgrounds; (d) was mainly produced in the United States, United Kingdom, and Germany; (d) has fragmented research networks and a limited number of researchers per network; (e) has limited cross-country collaboration except for that between the US and the UK in one hand and countries in the Mekong region in the other hand; (f) emphasized on medications related to malaria and sexual stimulants; and (g) received relatively inadequate funding.
CONCLUSIONS
Research on SF medical products is important and should remain a priority to ensure good quality of medications. Research activity in the field needs to be encouraged in world regions such as Africa and the Middle East where drug regulations are unsatisfactory and cross-border trade of illegal medications is common.
Topics: Bibliometrics; Delivery of Health Care; Drug and Narcotic Control; Humans; Public Health; Publications
PubMed: 34556126
DOI: 10.1186/s12992-021-00766-5 -
Genes & Development Oct 2021Double-strand break (DSB) repair choice is greatly influenced by the initial processing of DNA ends. 53BP1 limits the formation of recombinogenic single-strand DNA...
Double-strand break (DSB) repair choice is greatly influenced by the initial processing of DNA ends. 53BP1 limits the formation of recombinogenic single-strand DNA (ssDNA) in BRCA1-deficient cells, leading to defects in homologous recombination (HR). However, the exact mechanisms by which 53BP1 inhibits DSB resection remain unclear. Previous studies have identified two potential pathways: protection against DNA2/EXO1 exonucleases presumably through the Shieldin (SHLD) complex binding to ssDNA, and localized DNA synthesis through the CTC1-STN1-TEN1 (CST) and DNA polymerase α (Polα) to counteract resection. Using a combinatorial approach of END-seq, SAR-seq, and RPA ChIP-seq, we directly assessed the extent of resection, DNA synthesis, and ssDNA, respectively, at restriction enzyme-induced DSBs. We show that, in the presence of 53BP1, Polα-dependent DNA synthesis reduces the fraction of resected DSBs and the resection lengths in G0/G1, supporting a previous model that fill-in synthesis can limit the extent of resection. However, in the absence of 53BP1, Polα activity is sustained on ssDNA yet does not substantially counter resection. In contrast, EXO1 nuclease activity is essential for hyperresection in the absence of 53BP1. Thus, Polα-mediated fill-in partially limits resection in the presence of 53BP1 but cannot counter extensive hyperresection due to the loss of 53BP1 exonuclease blockade. These data provide the first nucleotide mapping of DNA synthesis at resected DSBs and provide insight into the relationship between fill-in polymerases and resection exonucleases.
Topics: DNA Breaks, Double-Stranded; DNA Repair; DNA Replication; DNA, Single-Stranded; Homologous Recombination; Tumor Suppressor p53-Binding Protein 1
PubMed: 34503990
DOI: 10.1101/gad.348667.121